植物脂肪酸的生物合成与基因工程

植物脂肪酸既具重要生理功能,又有巨大食用和工业价值。其生物合成途径较为复杂,涉及乙酰_CoA羧化酶、脂肪酸合成酶、脂肪酸去饱和酶和脂肪酸延长酶等一系列酶。近年来,对脂肪酸生物合成途径进行了大量研究,克隆出许多相关基因,初步阐明了脂肪酸合成规律,并在此基础上开展了利用基因工程技术调控脂肪酸合成研究,取得可喜进展。本文详细介绍了植物饱和脂肪酸、不饱和脂肪酸和超长链脂肪酸的生物合成与基因工程研究的新结果。     

植物脂肪酸的生物合成与基因工程

植物脂肪酸既具重要生理功能,又有巨大食用和工业价值。其生物合成途径较为复杂,涉及乙酰－ＣｏＡ羟化酶、脂肪酸合成酶、脂肪酸去饱和酶和脂肪酸延长酶等一系列酶。近年来,对脂肪酸生物合成途径进行了大量研究,克隆出许多相关基因,初步阐明了脂肪酸合成规律,并在此基础上开展了利用基因工程技术调控脂肪酸合成研究,取得可喜进展。本文详细介绍了植物饱和脂肪酸、不饱和脂肪酸和超长链脂肪酸的生物合成与基因工程研究的新结果     

Fatty acid transport across the cell membrane: Regulation by fatty acid transporters

Transport of long-chain fatty acids across the cell membrane has long been thought to occur by passive diffusion. However, in recent years there has been a fundamental shift in understanding, and it is now generally recognized that fatty acids cross the cell membrane via a protein-mediated mechanism. Membrane-associated fatty acid-binding proteins (‘fatty acid transporters’) not only facilitate but also regulate cellular fatty acid uptake, for instance through their inducible rapid (and reversible) translocation from intracellular storage pools to the cell membrane. A number of fatty acid transporters have been identified, including CD36, plasma membrane-associated fatty acid-binding protein (FABP_pm), and a family of fatty acid transport proteins (FATP1–6). Fatty acid transporters are also implicated in metabolic disease, such as insulin resistance and type-2 diabetes. In this report we briefly review current understanding of the mechanism of transmembrane fatty acid transport, and the function of fatty acid transporters in healthy cardiac and skeletal muscle, and in insulin resistance/type-2 diabetes. Fatty acid transporters hold promise as a future target to rectify lipid fluxes in the body and regain metabolic homeostasis.     

Fatty acyl-AMP as an intermediate in fatty acid reduction to aldehyde in luminescent bacteria

The acyl protein synthetase component (50K) of the fatty acid reductase complex from the luminescent system of Photobacterium phosphoreum has been found to catalyze the activation of fatty acid via formation of an enzyme bound acyl-AMP (carboxyphosphate mixed anhydride) immediately prior to the acylation of the enzyme. PPi-ATP exchange and nucleotide binding experiments are dependent on fatty acid and indicate that the fatty acyl-AMP is directly formed and that an adenylated enzyme intermediate is not part of the mechanism. The formation of acyl-AMP from fatty acid and ATP is reversible with a standard free energy of -2 kcal/mol, and is dependent on Mg2+. The fatty acyl-AMP intermediate has been isolated and shown to be part of the pathway of fatty acid reduction. The 34K component of the complex, which strongly stimulates the acylation of the 50K protein by fatty acyl-AMP or fatty acid and ATP, is not required for the formation of acyl-AMP showing that it differentially affects the fatty acid activation and acylation steps catalyzed by the 50K protein.     

侧柏叶子与种子脂肪酸的GC-MS比较分析研究

目的:研究柏子仁与侧柏叶的脂肪酸组成.方法:用GC-MS方法对侧柏叶子与种子油进行定性鉴定和定量分析.结果:鉴定了柏子仁油中的8种脂肪酸,占脂溶性成分的93.56%;侧柏叶子油中12种脂肪酸,占脂溶性成分的93.39%.柏子仁饱和脂肪酸主要是十六烷酸(8.11%)、硬脂酸(6.08%);不饱和脂肪酸主要为亚油酸(24.59%)、亚麻酸(59.77%),占脂肪酸的83.14%.侧柏叶子饱和脂肪酸饱和脂肪酸主要为十六烷酸(14.70%)、乙酸(3.20%)、十七烷酸(2.50%);不饱和脂肪酸主要为二十二碳四烯酸(40.48%)、亚油酸(10.69%)、亚麻酸(17.62%).结论:柏子仁和侧柏叶均含有合理的脂肪酸组成.     

Mechanism of fatty acid synthesis

Significant advances have been made in the past few years in our understanding of the mechanism of synthesis of fatty acids, the structural organization of fatty acid synthetase complexes and the mechanism of regulation of activity of these enzyme systems. Numerous fatty acid synthetase complexes have been purified to homogeneity and the mechanism of synthesis of fatty acids by these enzyme systems has been ascertained from tracer, and recently, kinetic studies. The results obtained by these methods are in complete agreement. Furthermore, the kinetic results have indicated that fatty acid synthesis proceeds by a seven-site ping-pong mechanism. Several of the fatty acid synthetases have been dissociated completely to nonidentical half-molecular weight subunit species and these have been separated by affinity chromatography. From one of these subunits acyl carrier protein has been obtained. Whether the nonidentical subunits can be dissociated into individual proteins or whether these subunits are each comprised of one peptide is still a matter of controversy. However, it appears to us that each of the half-molecular weight subunits is indeed comprised of individual proteins. Studies on the regulation of activity of fatty acid synthetase complexes of avian and mammalian liver have resulted in the separation by affinity chromatography of three species (apo, holo-$\text{a}$ and holo-$\text{b}$) of fatty acid synthetase. Since these species have radically different enzyme activities they may provide a mechanism of short-term regulation of fatty acid synthetase activity. Other studies have shown that the quantity of avian and mammalian liver fatty acid synthetases is controlled by a change in the rate of synthesis of this enzyme complex. This change in the rate of synthesis of enzyme complex is under the control of insulin and glucagon. The former hormone increases the rate of enzyme synthesis, whereas the latter decreases it. Further studies on fatty acid synthetase complexes will undoubtedly concentrate upon more refined aspects of the structural organization of these enzyme systems, including the sequencing of acyl carrier proteins, the effects of protein-protein interaction on the kinetics of the partial reactions of fatty acid synthesis catalyzed by separated enzymes of the complex, the mechanism of hormonal regulation of fatty acid synthetase activity and x-ray diffraction analysis of subunits and complex.     

Phytanic acid alpha-oxidation,new insights into an old problem: a review

Phytanic acid (3,7,10,14-tetramethylhexadecanoic acid) is a branched-chain fatty acid which is known to accumulate in a number of different genetic diseases including Refsum disease. Due to the presence of a methyl-group at the 3-position, phytanic acid and other 3-methyl fatty acids can not undergo beta-oxidation but are first subjected to fatty acid alpha-oxidation in which the terminal carboxyl-group is released as CO(2). The mechanism of alpha-oxidation has long remained obscure but has been resolved in recent years. Furthermore, peroxisomes have been found to play an indispensable role in fatty acid alpha-oxidation, and the complete alpha-oxidation machinery is probably localized in peroxisomes. This Review describes the current state of knowledge about fatty acid alpha-oxidation in mammals with particular emphasis on the mechanism involved and the enzymology of the pathway.     

Regulation of fatty acid transport

PURPOSE OF REVIEW: The aim of the present review is to summarize recent developments in the area of regulation of fatty acid transport. RECENT FINDINGS: While controversy still exists regarding the contribution of passive diffusion versus protein-mediated fatty acid transport, both processes are now widely accepted. With the recent identification of an increasing number of putative fatty acid transporters, emphasis has been placed on regulation including fatty acid transport function of the protein, and also possible associated functions (acylCoA synthase activity and vectorial channelling to intracellular processing). Deciphering these issues has been facilitated through the use of loss-of-function (such as knockout) and gain-of-function (cell transfectants and transgenic mice) models. SUMMARY: It is likely that our concept of fatty acid transport will continue to converge, incorporating the individual functions of the wide variety of fatty acid transporters into an integrated physiologic framework with relevance to a number of diseases.     

The fatty acid transport function of fatty acid-binding proteins

The intracellular fatty acid-binding proteins (FABPs) comprise a family of 14-15 kDa proteins which bind long-chain fatty acids. A role for FABPs in fatty acid transport has been hypothesized for several decades, and the accumulated indirect and correlative evidence is largely supportive of this proposed function. In recent years, a number of experimental approaches which more directly examine the transport function of FABPs have been taken. These include molecular level in vitro modeling of fatty acid transfer mechanisms, whole cell studies of fatty acid uptake and intracellular transfer following genetic manipulation of FABP type and amount, and an examination of cells and tissues from animals engineered to lack expression of specific FABPs. Collectively, data from these studies have provided strong support for defining the FABPs as fatty acid transport proteins. Further studies are necessary to elucidate the fundamental mechanisms by which cellular fatty acid trafficking is modulated by the FABPs.     

Fatty acids covalently bound to erythrocyte proteins undergo a differential turnover in vivo

Recently, covalently bound fatty acids have been identified on a variety of proteins. Many of these acyl proteins are physiologically important, and the lipid modification often appears to be essential for their function. In this investigation mature erythrocytes have been used to study in detail the metabolic behavior of protein-bound fatty acids. Although deficient in protein synthesis, these cells are still able to covalently attach [3H]palmitic acid to proteins located at the plasma membrane and its associated cytoskeleton. Linkage analyses demonstrated that the labeled polypeptides contained ester- or thioester-bound fatty acids. The covalent binding of fatty acid was rapidly reversible. Half-lives of the protein-bound fatty acid molecules ranged from less than 30 min to more than 3 h. The deacylation reaction was not due to a chemically labile linkage of protein and fatty acid but appeared to be physiologically induced. Differences in the fatty acid turnover rates between the acyl proteins suggested an independent regulation of their lipid turnover. A number of proteins underwent dynamic fatty acid acylation, indicating that palmitylated proteins undergoing fatty acid turnover are not a rare exception.     

Cytoplasmic fatty acid-binding proteins: emerging roles in metabolism and atherosclerosis

Cytoplasmic fatty acid-binding proteins (FABPs) are a family of proteins, expressed in a tissue-specific manner, that bind fatty acid ligands and are involved in shuttling fatty acids to cellular compartments, modulating intracellular lipid metabolism, and regulating gene expression. Several members of the FABP family have been shown to have important roles in regulating metabolism and have links to the development of insulin resistance and the metabolic syndrome. Recent studies demonstrate a role for intestinal FABP in the control of dietary fatty acid absorption and chylomicron secretion. Heart FABP is essential for normal myocardial fatty acid oxidation and modulates fatty acid uptake in skeletal muscle. Liver FABP is directly involved in fatty acid ligand signaling to the nucleus and interacts with peroxisome proliferator-activated receptors in hepatocytes. The adipocyte FABP (aP2) has been shown to affect insulin sensitivity, lipid metabolism and lipolysis, and has recently been shown to play an important role in atherosclerosis. Interestingly, expression of aP2 by the macrophage promotes atherogenesis, thus providing a link between insulin resistance, intracellular fatty acid disposition, and foam cell formation. The FABPs are promising targets for the treatment of dyslipidemia, insulin resistance, and atherosclerosis in humans.     

Cellular fatty acid uptake: the contribution of metabolism

PURPOSE OF REVIEW: The aim of this review is to highlight the importance of fatty acid metabolism as a major determinant in fatty acid uptake. In particular, we emphasize how the activation, intracellular transport and downstream metabolism of fatty acids influence their uptake into cells. RECENT FINDINGS: Studies examining fatty acid entry into cells have focused primarily on the roles of plasma membrane proteins or the question of passive diffusion. Recent studies, however, strongly suggest that a driving force governing fatty acid uptake is the metabolic demand for fatty acids. Both gain and loss-of-function experiments indicate that fatty acid uptake can be modulated by activation at both the plasma membrane and internal sites, by intracellular fatty acid binding proteins, and by enzymes in synthetic or degradative metabolic pathways. Although the mechanism is not known, it appears that converting fatty acids to acyl-CoAs and downstream metabolic intermediates increases cellular fatty acid uptake, probably by limiting efflux. SUMMARY: Altered fatty acid metabolism and the accumulation of triacylglycerol and lipid metabolites has been strongly associated with insulin resistance and diabetes, but we do not fully understand how the entry of fatty acids into cells is regulated. Future studies of cellular fatty acid uptake should consider the influence of fatty acid metabolism and the possible interactions between fatty acid metabolism or metabolites and fatty acid transport proteins.     

Distribution and fatty acid composition of phosphoglycerides in normal human brain

A thin-layer chromatographic procedure for the isolation of tissue phospholipids and their subsequent analysis is described. The method has been applied to the determination of the fatty acids of phosphoglycerides in human brain from the early fetal stage to old age. The study shows changes in the distribution and fatty acid composition of each phosphoglyceride in normal brain, although they are quite small after early childhood. A lipid-specific fatty acid pattern for each of the four major phosphoglycerides was found. Besides this, the pronounced differences between fatty acids of the lipids from the cerebral cortex and from the adjacent white matter justify speaking of a tissue-specific fatty acid pattern for brain phosphoglycerides. The phospholipids of cerebral white matter contained more monoenoic acid but much less polyunsaturated fatty acid than those of cerebral cortex. The brain phosphoglycerides also showed an age-dependent fatty acid pattern. With increasing age the concentration of the fatty acids of the linoleate family diminished while that of the linolenate family increased. Brain inositol phosphoglycerides, the fatty acid composition of which has not been studied systematically before, were characterized by a large concentration of arachidonate which was nearly as high for white as for gray matter and showed only small changes with age.     

Composition of phospholipids and of phospholipid fatty acids and aldehydes in human red cells

Improved methods for lipid analysis that have been developed recently were employed to reevaluate the phospholipid composition, the fatty acid and fatty aldehyde composition of the total phospholipid, and the fatty acid composition of the individual phospholipids of normal human red cells. Thirty-three fatty acids and five fatty aldehydes were estimated and tentatively identified in the total phospholipid of normal human red cells. Additional minor components were evident. The major individual phospholipids were isolated by silicic acid thin-layer chromatography and quantified. The fatty acid compositions of phosphatidyl ethanolamine, phosphatidyl serine, lecithin, and sphingomyelin were determined. Each of these phospholipids showed a distinctive and characteristic fatty acid pattern.     

Membrane proteins implicated in long-chain fatty acid uptake by mammalian cells: CD36, FATP and FABPm

Long-chain fatty acids can transfer passively across mammalian cell membranes. However, under physiological conditions of low fatty acid to albumin ratios in the circulation, the major fraction of uptake appears to be mediated by a saturable, protein-facilitated component. A simple diffusion process becomes significant at high molar ratios of fatty acid to albumin as the concentration of free fatty acid in solution is increased. Identification of the mammalian membrane fatty acid transporter(s) has been the focus of active investigation by several research groups. In this review we discuss three candidate proteins: FABPm, FAT/CD36 and FATP which have been cloned and are currently being characterized. Recent evidence arguing for an important role of the fatty acid transport step in general metabolism and linking these proteins to physiologic or metabolic abnormalities is described.     

Relationship between fatty acid binding proteins,acetyl-CoA formation and fatty acid synthesis in developing human placenta

The relationship between fatty acid binding proteins, ATP citrate lyase activity and fatty acid synthesis in developing human placenta has been studied. Fatty acid binding proteins reverse the inhibitory efect of palmitoyl-CoA and oleate on ATP citrate lyase and fatty acid synthesis. In the absence of these inhibitors fatty acid binding proteins activate ATP citrate lyase and stimulate [ 1-¹⁴ C] acetate incorporation into placental fatty acids indicating binding of endogenous inhibitors by these proteins. Thus these proteins regulate the supply of acetyl-CoA as well as the synthesis of fatty acids from that substrates. As gestation proceeds and more lipids are required by the developing placenta fatty acid binding protein content, activity of ATP citrate lyase and rate of fatty acid synthesis increase indicating a cause and efect relationship between the demand of lipids and supply of precursor fatty acids during human placental development.     

Formation of alpha-tocopherol complexes with fatty acids. Nature of complexes

Using ultraviolet spectrophotometry and 1H-NMR high-resolution spectroscopy, it has been demonstrated that the formation of alpha-tocopherol complexes with free fatty acids occurs via two types of interaction, namely formation of a hydrogen bond between the alpha-tocopherol chromanol nucleus hydroxyl and the carboxyl group of a fatty acid, and interaction of the fatty acid acyl chains with the chromanol nucleus methyl groups. The second interaction is significantly enhanced by an increase in the number of double bonds in the fatty acid molecule, which results in restriction of the molecular mobility of alpha-tocopherol. The proposed structural model of alpha-tocopherol-fatty acid complexes has been confirmed by the use of molecular models. It has been assumed that the efficiency of complex formation of natural tocopherols with fatty acids is correlated with their biological activity.     

On the mechanism of enzyme action. XLVIII. Investigation of the fat of Fusarium lini Bolley by means of urea adducts

The usefulness of urea adducts for the fractionation of natural fatty acids has been demonstrated. Stearic acid has been isolated and identified from the fat of Fusarium lini Bolley by purification of the solid fraction obtained from the urea adduct of its fatty acids. The conversion of stearic acid to fatty material with a higher iodine value has provided further evidence for the enzymatic formation of unsaturated fatty acids in Fusarium lini Bolley by means of the action of a fatty acid dehydrogenase system.     

Fatty acid alterations and n-3 fatty acid supplementation in cystic fibrosis

Specific fatty acid alterations have been described in the blood and tissues of cystic fibrosis (CF) patients. The two most consistent alterations include decreased levels of linoleic acid (LA) and decreased levels of docosahexaenoic acid (DHA). Increased arachidonic acid (AA) release from membrane phospholipids, as well as changes in levels of AA and other monounsaturated and polyunsaturated fatty acids (PUFAs) have also been described in CF. Although mechanisms of fatty acid alterations have not yet been determined, these alterations may have an important role in the progression of the CF disease. There have been several clinical trials in which CF patients were supplemented with n-3 fatty acids. Most trials resulted in an increase in the levels of the supplemental fatty acids in the blood of CF patients in the absence of significant clinical improvement. It is recommended that future trials include a larger population of CF patients and measure multiple clinical outcomes.     

Role of fatty acid binding protein in the modulation of inhibitory effect of fatty acids on fatty acid synthase and ATP-citrate lyase in developing human brain.

Inhibition of the activities of fatty acid synthase and ATP-citrate lyase (ATP-CL) by fatty acids and their CoA esters has been studied. Purified fatty acid binding protein from human fetal brain reverses this inhibition. This protein also activates the enzyme when added alone. ATP-citrate lyase and fatty acid synthase activity gradually increased with the advancement of gestation showing a relationship between high demand of fatty acid synthesis in developing brain and supply of its precursors.